Cosmetic compositions

ABSTRACT

A topical composition having (i) an isolated polypeptide having the sequence of SEQ ID NO: 1 or a functional equivalent thereof, and (ii) a cosmetically acceptable carrier; and use thereof.

BACKGROUND

Aging is a natural process not yet totally understood. It appears to begenetically controlled and can be accelerated by environmental, dietary,and pathological factors. At the cellular or molecular level, aging isconsidered as the result of the inability of an organism to respondadaptively to environmental changes. As people age, the process ofself-renewal or replacement of skin cells slows down.

The skin has three layers: the epidermis, the dermis, and thehypodermis. The epidermis, the outmost layer, is filled with strata ofspecialized skin cells known as keratinocytes. These cells start out inthe deepest layer of the epidermis and migrate to the skin surface.During this process, they lose moisture and are eventually sloughed off.In young people, this process takes about 4 weeks to travel to thesurface. In people of age over 50, it can take about 5.5 weeks. Thedermis, the layer beneath the epidermis, contains blood vessels thatnourish the skin cells and the structural elements e.g., collagen andelastin, which keep the skin firm and springy. Aging results in fewerkeratinocyte layers and, thus, less skin stem cell self-renewal andreplacement. Also, the aged skin lost collagen and elastin poorly,leading to thin and papery-look appearance. Consequently, wrinkles form.This slow, concurrent losses of dermal cellular and structural elements,thickness, and elasticity, together with wrinkle and segment formation,are the most obvious indicators of skin aging.

Various compositions and methods have been used for regulatingundesirable skin surface texture, such as fine lines, wrinkles, pores,and the like. However, many of them are ineffective or cause allergy andother side effects. Thus, there is a need for a more effective and safercosmetic composition.

SUMMARY

This invention relates to cosmetic compositions and use thereof.Accordingly, one aspect of this invention features a topical compositioncontaining an isolated polypeptide having the sequence ofstromal-derived factor-1 (SDF-1) variant named stem cell active factor(SCAF) or its functional equivalent, and a cosmetically acceptablecarrier. Shown below are the polypeptide sequence of SCAF and its cDNAsequence (SEQ ID NOs: 1 and 2, respectively)

(SEQ ID NO: 1) MNAKVVVVLVLVLTALCLSDGKPVSLSYRCPCRFFESHVARANVKHLKILNTPNCALQIVARLKNNNRQVSIDPKLKWIQEYLEKALNK (SEQ ID NO: 2)ATGAACGCCAAGGTCGTGGTCGTGCTGGTCCTCGTGCTGACCGCGCTCTGCCTCAGCGACGGGAAGCCCGTCAGCCTGAGCTACAGATGCCCATGCCGATTCTTCGAAAGCCATGTTGCCAGAGCCAACGTCAAGCATCTCAAAATTCTCAACACTCCAAACTGTGCCCTTCAGATTGTAGCCCGGCTGAAGAACAACAACAGACAAGTGTCCATTGACCCGAAGCTAAAGTGGATTCAGGAGTACCTGG AGAAAGCTTTAAACAAGTAA

An “isolated polypeptide” refers to a polypeptide that has beenseparated from other proteins, lipids, and nucleic acids with which itis naturally associated. The polypeptide can constitutes at least 10%(i.e., any percentage between 10% and 100%) by dry weight of thepurified preparation. Purity can be measured by any appropriate standardmethod, for example, by column chromatography, polyacrylamide gelelectrophoresis, or HPLC analysis. An isolated polypeptide of theinvention can be purified from a natural source, produced by recombinantDNA techniques, or by chemical methods. A “functional equivalent” refersto a polypeptide derivative of the SCAF polypeptide, e.g., a proteinhaving one or more point mutations, insertions, deletions, truncations,a fusion protein, or a combination thereof. It retains substantially theactivity of the SCAF polypeptide, i.e., the ability to improve skincondition (e.g., reducing wrinkles). The isolated polypeptide cancontain SEQ ID NO: 1 or a functional fragment of SEQ ID NO: 1. Ingeneral, the functional equivalent is at least 75% (e.g., any numberbetween 75% and 100%, inclusive) identical to SEQ ID NO: 1. A“cosmetically acceptable” or “dermatologically-acceptable” compositionor component refers a composition or component that is suitable for usein contact with human skin without undue toxicity, incompatibility,instability, allergic response, and the like.

The above-described topical composition can further contain an agentselected from the group consisting of an anti-wrinkle herbal extract, awhiting herbal extract, and an anti-acne herbal extract. Examples of theanti-wrinkle herbal extract include a hop extract, a horsetail extract,or both. The whiting herbal extract can contain a Scutellariabaicalensis root extract, a Saxifrage sarmentosa root extract, or both.Examples of the anti-acne herbal extract includes a watercress extract,a witch hazel extract, a centella extract, a licorice extract, or acombination thereof. The topical composition can further contain a purepolysaccharide, e.g., L-fucose, D-galactose, galacturonic acid, or acombination thereof.

The topical composition of the invention provides an essentiallyimmediate improvement in skin feel and appearance. It is also useful forproviding visual improvements in skin appearance or conditions followingmultiple topical applications. Further, the composition provides thevisual benefits without imparting an unacceptable skin appearance.

Thus, the invention also features a method of reducing skin wrinkles orpromoting hair growth. The method includes applying to a surface of skinin need thereof a safe and effective amount of the above-describedtopical composition. A safe and effective amount refers to an amount ofa compound, component, or composition sufficient to significantly inducea positive benefit, preferably a positive skin appearance or feelbenefit, including independently the benefits disclosed herein, but lowenough to avoid serious side effects, i.e., to provide a reasonablebenefit to risk ratio, within the scope of sound medical judgment.

The invention features an isolated polypeptide containing theabove-listed SEQ ID NO: 1 and an isolated nucleic acid that contains asequence encoding the polypeptide, e.g., SEQ ID NO: 2. The polypeptidecan be used in the composition described above

A nucleic acid refers to a DNA molecule (e.g., a cDNA or genomic DNA),an RNA molecule (e.g., an mRNA), or a DNA or RNA analog. A DNA or RNAanalog can be synthesized from nucleotide analogs. The nucleic acidmolecule can be single-stranded or double-stranded, but preferably isdouble-stranded DNA. An “isolated nucleic acid” is a nucleic acid thestructure of which is not identical to that of any naturally occurringnucleic acid or to that of any fragment of a naturally occurring genomicnucleic acid. The term therefore covers, for example, (a) a DNA whichhas the sequence of part of a naturally occurring genomic DNA moleculebut is not flanked by both of the coding sequences that flank that partof the molecule in the genome of the organism in which it naturallyoccurs; (b) a nucleic acid incorporated into a vector or into thegenomic DNA of a prokaryote or eukaryote in a manner such that theresulting molecule is not identical to any naturally occurring vector orgenomic DNA; (c) a separate molecule such as a cDNA, a genomic fragment,a fragment produced by polymerase chain reaction (PCR), or a restrictionfragment; and (d) a recombinant nucleotide sequence that is part of ahybrid gene, i.e., a gene encoding a fusion protein. The nucleic aciddescribed above can be used to express the polypeptide of thisinvention. For this purpose, one can operatively linked the nucleic acidto suitable regulatory sequences to generate an expression vector.

A vector refers to a nucleic acid molecule capable of transportinganother nucleic acid to which it has been linked. The vector can becapable of autonomous replication or integrate into a host DNA. Examplesof the vector include a plasmid, cosmid, or viral vector. The vector ofthis invention includes a nucleic acid in a form suitable for expressionof the nucleic acid in a host cell. Preferably the vector includes oneor more regulatory sequences operatively linked to the nucleic acidsequence to be expressed. A “regulatory sequence” includes promoters,enhancers, and other expression control elements (e.g., polyadenylationsignals). Regulatory sequences include those that direct constitutiveexpression of a nucleotide sequence, as well as tissue-specificregulatory and/or inducible sequences. The design of the expressionvector can depend on such factors as the choice of the host cell to betransformed, the level of expression of protein desired, and the like.The expression vector can be introduced into host cells to produce thepolypeptide of this invention. Also within the scope of this inventionis a host cell that contains the above-described nucleic acid. Examplesinclude E. coli cells, insect cells (e.g., using baculovirus expressionvectors), yeast cells, or mammalian cells. See e.g., Goeddel, (1990)Gene Expression Technology: Methods in Enzymology 185, Academic Press,San Diego, Calif. To produce a polypeptide of this invention, one canculture a host cell in a medium under conditions permitting expressionof the polypeptide encoded by a nucleic acid of this invention, andpurify the polypeptide from the cultured cell or the medium of the cell.Alternatively, the nucleic acid of this invention can be transcribed andtranslated in vitro, for example, using T7 promoter regulatory sequencesand T7 polymerase.

The details of one or more embodiments of the invention are set forth inthe description below. Other features, objects, and advantages of theinvention will be apparent from the description and from the claims.

DETAILED DESCRIPTION

This invention is based, at least in part, on unexpected findings that atopical composition containing SCAF regulates and improves aging skinconditions. As mentioned above, the principal process of aging takeplace both in the top layers of skin (i.e. the epidermis) and in thedermis. A composition of this invention not only regulates and improvesthe conditions of the epidermis, but also effects the dermis byimproving the nourishment and activating stem cells to renew anddifferentiate into fibroblasts, keratinocytes, and other cellularcomponents of the skin.

SCAF, an SDF-1 variant, described herein is a member of a chemokinefamily consisting of small secreted proteins (8-12 kDa). This family ofchemokines are known to cause activation and migration of leukocytes(Baggiolini, 1998, Nature 392, 565-8; and Murdoch et al., 2000, Blood95, 3032-43). As described in the examples below, a composition of thisinvention unexpectedly improves the condition of the epidermis layer orhair growth, without causing side effects related to leukocyteactivation and migration, such as allergy. Thus, SCAF and other SDF-1variants can be used as an active ingredient in a cosmetic composition.

While many SDF-1 preparations can be used, highly purified SDF-1 ispreferred. Examples of SDF-1 include mammalian SDF-1 (e.g., human SDF-1)or SDF-1 having substantially the same biological activity as mammalianSDF-1. All of naturally occurring SDF-1, genetic engineered SDF-1, andchemically synthesized SDF-1 can be used. SDF-1 obtained by recombinantDNA technology may have the same amino acid sequence as naturally aoccurring SDF-1 (SEQ ID NO: 1) or an functionally equivalent thereof.The term “SDF-1” also covers chemically modified SDF-1. Examples ofchemically modified SDF-1 include SDF-1 subjected to conformationalchange, addition or deletion of a sugar chain, and SDF-1 to which acompound such as polyethylene glycol has been bound. Once purified andtested by standard methods or according to the method described in theexamples below, SDF-1 can be included in a topical composition.

The amino acid composition of the SDF-1 polypeptide described herein mayvary without disrupting the ability of the polypeptide to improve skincondition. For example, it can contain one or more conservative aminoacid substitutions. A “conservative amino acid substitution” is one inwhich the amino acid residue is replaced with an amino acid residuehaving a similar side chain. Families of amino acid residues havingsimilar side chains have been defined in the art. These families includeamino acids with basic side chains (e.g., lysine, arginine, histidine),acidic side chains (e.g., aspartic acid, glutamic acid), uncharged polarside chains (e.g., glycine, asparagine, glutamine, serine, threonine,tyrosine, cysteine), nonpolar side chains (e.g., alanine, valine,leucine, isoleucine, proline, phenylalanine, methionine, tryptophan),beta-branched side chains (e.g., threonine, valine, isoleucine) andaromatic side chains (e.g., tyrosine, phenylalanine, tryptophan,histidine). Thus, a predicted nonessential amino acid residue in SEQ IDNO: 1 is preferably replaced with another amino acid residue from thesame side chain family. Alternatively, mutations can be introducedrandomly along all or part of SEQ ID NO: 1, such as by saturationmutagenesis, and the resultant mutants can be screened for the abilityto improve skin condition to identify mutants that retain the activityas descried below in Example 3.

A polypeptide of the invention can be obtained as a syntheticpolypeptide or a recombinant polypeptide. To prepare a recombinantpolypeptide, a nucleic acid encoding it can be linked to another nucleicacid encoding a fusion partner, e.g., glutathione-s-transferase (GST),6×-His epitope tag, or M13 Gene 3 protein. The resultant fusion nucleicacid expresses in suitable host cells a fusion protein that can beisolated by methods known in the art. The isolated fusion protein can befurther treated, e.g., by enzymatic digestion, to remove the fusionpartner and obtain the recombinant polypeptide of this invention.

The topical composition of this invention can further contain ananti-wrinkle herbal extract, a whiting herbal extract, or an anti-acneherbal extract. An extract is a concentrated preparation of theessential constituents of a medicinal herb. Typically, the essentialconstituents are extracted from a herb by suspending the herb in anappropriate choice of solvent, typically, water, ethanol/water mixture,methanol, butanol, iso-butanol, acetone, hexane, petroleum ether orother suitable organic solvents. The extracting process may be furtherfacilitated by means of maceration, percolation, repercolation,counter-current extraction, turbo-extraction, or by carbon-dioxidehypercritical (temperature/pressure) extraction. After filtration to ridof herb debris, the extracting solution may be further evaporated andthus concentrated to yield a soft extract (extractum spissum) oreventually a dried extract (extracum siccum), by means of spray drying,vacuum oven drying, fluid-bed drying, or freeze-drying. The soft extractor dried extract may be further dissolved in a suitable liquid to adesired concentration for administering or processed into a suitableform.

In a 100 g topical composition of this invention, exemplary quantitiesof the herb extract ingredients are: the anti-wrinkle herbal extract3-30 g (e.g., 15 g); the whiting herbal extract, 3-30 g (e.g., 15 g);the anti-acne herbal extract: 3-30 g (e.g., 15 g); and SDF-1 2000-5000Unit (e.g., 3000 unit). One “Unit” is defined as the amount of SDF-1that, after administered to a subject, induce the migration of 1×10⁶peripheral blood cells for at least 3 mm in vitro. The anti-wrinkleherbal extract can be prepared from hop or horsetail; the whiting herbalextract can be obtained from Scutellaria baicalensis root, or Saxifragesarmentosa root; and the anti-acne herbal extract can be obtained fromwatercress, witch hazel, centella extract, or licorice. These herbs arecommercially available. After authenticating each herb, conventionalmethods may be used to process the composition of the present inventioninto a form suitable for administering to human subjects. Those methodsare either described in pertinent literature or commonly used bypractitioners of herbal medicine.

The composition of the present invention also contains a safe andeffective amount of a dermatologically acceptable carrier that issuitable for topical application to the skin. It enables the essentialmaterials and optional components in it to be delivered to the skin atan appropriate concentration. The carrier can thus act as a diluent,dispersant, solvent, or the like to ensure that the active materials areapplied to and distributed evenly over the selected target at anappropriate concentration. The carrier can be solid, semi-solid, orliquid. Preferably, the carrier is in the form of a lotion, a cream, ora gel, more preferably one that has a sufficient thickness or yieldpoint to prevent the active materials from sedimenting. The carrier canitself be inert or it can possess dermatological benefits of its own.The carrier should also be physically and, chemically compatible withthe essential components described herein, and should not unduly impairstability, efficacy, or other use benefits associated with thecompositions of the present invention.

The type of carrier utilized in the present invention depends on thetype of product form desired for the composition. The topicalcompositions useful in the subject invention may be made into a widevariety of product forms such as are known in the art. These include,but are not limited to, lotions, creams, gels, sticks, sprays,ointments, pastes, and mousses. These product forms may comprise severaltypes of carriers including, but not limited to, solutions, aerosols,emulsions, gels, solids, and liposomes.

Preferred carriers can contain a dermatologically acceptable,hydrophilic diluent. Suitable hydrophilic diluents include water,organic hydrophilic diluents, such as C₁-C₄ monohydric alcohols and lowmolecular weight glycols and polyols (including propylene glycol,polyethylene glycol of, e.g., MW 200-600), polypropylene glycol of, e.g.MW 425-2025, glycerol, butylene glycol, 1,2,4-butanetriol, sorbitolesters, 1,2,6-hexanetriol, ethanol, iso-propanol, sorbitol esters,ethoxylated ethers, propoxylated ethers, and combinations thereof. Thecomposition preferably comprises at least about 60% of the hydrophilicdiluent.

Preferred carriers also contain an emulsion having a hydrophilic phase,especially an aqueous phase, and a hydrophobic phase e.g., a lipid, oil,or oily material. As well known to one skilled in the art, thehydrophilic phase will be dispersed in the hydrophobic phase, or viceversa, to form respectively hydrophilic or hydrophobic dispersed andcontinuous phases, depending on the composition ingredients. The term“dispersed phase,” a term well-known to one skilled in the art, refersto a phase that exists as small particles or droplets suspended in andsurrounded by a continuous phase. The dispersed phase is also known asthe internal or discontinuous phase. The emulsion may be or contain(e.g., in a triple or other multi-phase emulsion) an oil-in-wateremulsion or a water-in-oil emulsion such as a water-in-siliconeemulsion. Oil-in-water emulsions typically comprise from about 1% toabout 50% (preferably about 1% to about 30%) of the dispersedhydrophobic phase and from about 1% to about 99% (preferably from about40% to about 90%) of the continuous hydrophilic phase; water-in-oilemulsions typically comprise from about 1% to about 98% (preferably fromabout 40% to about 90%) of the dispersed hydrophilic phase and fromabout 1% to about 50% (preferably about 1% to about 30%) of thecontinuous hydrophobic phase. The emulsion may also comprise a gelnetwork, such as described in G. M. Eccleston, Application of EmulsionStability Theories to Mobile and Semisolid O/W Emulsions, Cosmetics &Toiletries, Vol. 101, November 1996, pp. 73-92, incorporated herein byreference. Preferred compositions herein are oil-in-water emulsions.

Preferred examples of the topical composition of this invention have anapparent viscosity of from about 5,000 to about 200,000 mPa·s(centipoise). For example, preferred lotions have an apparent viscosityof from about 10,000 to about 40,000 mPa·s; preferred creams have anapparent viscosity of from about 30,000 to about 160,000 mPa·s. Apparentviscosity can be determined using a Brookfield DVII RV viscometer,spindle TD, at 5 rpm, or the equivalent thereof. The viscosity isdetermined on a composition after the composition has been allowed tostabilize following its preparation, generally at least 24 hours underconditions of 25° C.±1° C. and ambient pressure after preparation of thecomposition. Apparent viscosity is measured with the composition at atemperature of 25° C.±1° C., after 30 seconds spindle rotation.

The topical composition of the present invention is usually formulatedto have a pH of 9.5 or below and in general have a pH in the range fromabout 4.5 to about 9, more preferably from about 5 to about 8.5. Someexamples, particularly those containing an additional active agent suchas salicylic acid, require a lower pH in order for the additional activeto be fully efficacious. These compositions are usually formulated tohave a pH of from about 2.5 to about 5, more preferably from about 2.7to about 4.

The topical compositions may contain a wide variety of optionalcomponents, provided that such optional components are physically andchemically compatible with the essential components described herein,and do not unduly impair stability, efficacy, or other use benefitsassociated with the compositions. Optional components may be dispersed,dissolved, or the like in the carrier of the present compositions.

Exemplary optional components include emollients, oil absorbents,antimicrobial agents, binders, buffering agents, denaturants, cosmeticastringents, external analgesics, film formers, humectants, opacifyingagents, perfumes, pigments, skin soothing and healing agents,preservatives, propellants, skin penetration enhancers, solvents,suspending agents, emulsifiers, cleansing agents, thickening agents,solubilising agents, waxes, sunscreens, sunless tanning agents,antioxidants and/or radical scavengers, chelating agents, anti-acneagents, anti-inflammatory agents, desquamation agents/exfoliants,organic hydroxy acids, vitamins, and natural extracts. Examples of suchmaterials are described in Harry's Cosmeticology, 7th Ed., Harry &Wilkinson (Hill Publishers, London 1982); in Pharmaceutical DosageForms—Disperse Systems; Lieberman, Rieger & Banker, Vols. 1 (1988) & 2(1989); Marcel Decker, Inc.; in The Chemistry and Manufacture ofCosmetics, 2nd. Ed., deNavarre (Van Nostrand 1962-1965); and in TheHandbook of Cosmetic Science and Technology, 1st Ed. Knowlton & Pearce(Elsevier 1993) can also be used in the present invention.

The topical composition of the present invention is generally preparedby conventional methods such as are known in the art of making topicalcompositions. Such methods typically involve mixing of the ingredientsin one or more steps to a relatively uniform state, with or withoutheating, cooling, application of vacuum, and the like.

The topical composition is useful for regulating or improving skincondition, including regulating visible or tactile wrinkles ordiscontinuities in skin, e.g., visible and/or tactile wrinkles ordiscontinuities in skin texture or color, more especially thoseassociated with skin ageing. Such wrinkles or discontinuities may beinduced or caused by internal factors (e.g., chronological aging andother biochemical changes from within the skin) or external factors(e.g., ultraviolet radiation, environmental pollution, wind, heat, lowhumidity, harsh surfactants, and abrasives).

Regulating skin conditions can be carried out prophylactically ortherapeutically. Prophylactical regulation includes delaying,minimizing, or preventing visible or tactile wrinkles or discontinuitiesin skin. Therapeutic regulation, on the other hand, includesameliorating, diminishing, minimizing or effacing such wrinkles ordiscontinuities. Regulating skin conditions involves improving skinappearance feel, e.g., providing a smoother, more even appearance, orfeel and reducing signs of aging.

To use a topical composition of this invention, one can topically applyto the skin a safe and effective amount of the composition. The appliedamount, the frequency of application and the period of use vary widelydepending upon the active levels of a given composition and the level ofregulation desired, e.g., in light of the level of skin ageing presentin the subject and the rate of further skin ageing.

A wide range of quantities of the compositions of the present inventioncan be employed to provide a skin appearance and/or feel benefit.Quantities of the compositions typically applied per application arefrom about 0.1 mg/cm² to about 10 mg/cm², e.g., 2 mg/cm². Typically, acomposition can be used once per day. However application rates can varyfrom about once per week up to about three times per day or more.

The topical compositions of this invention provides a visibleimprovement in skin condition essentially immediately followingapplication of the composition to the skin. Such immediate improvementinvolves coverage or masking of skin imperfections such as texturaldiscontinuities (including those associated with skin aging, e.g.,enlarged pores), or providing a more even skin tone or color. Thecompositions of the invention also provide visible improvements in skincondition following chronic topical application. “Chronic topicalapplication” involves continued topical application of a compositionover an extended period during the subject's lifetime, preferably for aperiod of at least about one week, one month, three months, six months,or one year. Chronic regulation of skin condition involves improvementof skin condition following multiple topical applications.

Regulating skin conditions is preferably performed by applying acomposition in the form of a skin lotion, cream, cosmetic, or the likewhich is intended to be left on the skin for an extended period for someaesthetic, prophylactic, therapeutic or other benefit (i.e., a“leave-on” composition). As used herein, “leave-on” compositions excluderinse-off skin cleansing products. After applying the composition to theskin, the leave-on composition is preferably left on the skin for aperiod of at least about 15 minutes, 30 minutes, 1 hour, or up to about12 hours.

The specific examples below are to be construed as merely illustrative,and not limitative of the remainder of the disclosure in any waywhatsoever. Without further elaboration, it is believed that one skilledin the art can, based on the description herein, utilize the presentinvention to its fullest extent.

EXAMPLE 1

A number of extracts were prepared from herb plants. In general, freshplants were used to prepare extracts based on the principle defined inthe German Homeopathic Pharmacopoeia (HAB 1/78). Fresh plants should becollected or harvested in dry whether and be as free of dust or dirt aspossible. The plants must have no external sign of disease and must havenot withered or dead parts, major injuries, rot, or other changes notinherent in the species. If necessary, the plants should be washed withas little water as possible. The plants should then processedimmediately. Unless specified otherwise, the following harvesting timesshould be observed:

Harvested target Harvesting time Whole plants (including undergroundWhen in flower parts) Herbs (excluding underground parts After completedevelopment, and including leaves and shoots) shortly before or at startof blossoming Blossom Shortly after opening Barks Autumn or spring Rootsand root stocks Annuals, when seeds are ripe; perennials, spring Fruitsand seeds When ripe Unripe fruits Before ripening Mushrooms Aftercomplete development of fruiting bodyAfter harvesting, the plants or plant parts were cleaned, crushed, andextracted in a solvent, such as propylene glycol/ethanol or ethanol.

A hop extract was obtained using a solvent of water, propylene glycol,and ethanol (43%, 42%, and 15%, respectively). The resultant extract wasa brown-yellow liquid with an aromatic odor, a sweetish-bitter taste, apH value of 4.4-5.5, and a density of 1.000-1.0015 g/ml. It was solublein water, 60% ethanol, or 94% ethanol.

A horsetail extract was prepared from Equisetum arvense using thejust-mentioned solvent. The extract was a yellow liquid with asweetish-bitter taste, no odor, a pH value of 5.5-6.5, and a density of1.000-1.0015 g/ml. The extract was soluble in water and 60% ethanol, butformed a slightly opalized solution in 94% ethanol. Active ingredientsin this extract included silicic acid, saponin, and flavonoids.

A Scutellaria baicalensis root extract was prepared using ethanol fromthe root of Scutellaria baicalensis Georgi (labiatae). Beforeextracting, the periderm was removed in 50% 1,3-butylene glycol solutionby heating, refluxing, and filtering. The resulting filtrate wasconcentrated to 1/20 in volume and extracted with pure water. Themixture was then mixed with the same volume of 1,3-butylene glycol andfiltered. The filtrate retained was a yellowish brown clear liquid witha characteristic odor.

A Saxifrage sarmentosa root extract was prepared from Saxifragastolonifera Meerburg (Saxifragaceae) using 1,3-butylene glycol.Saxifrage sarmentosa roots were cut into pieces and mixed with 30 v/v %1,3-butylene glycol (120 litter for 10 kg Saxifrage sarmentosa). Theresultant mixture was filtered to obtain a clear, brown liquid, whichhad a pH value of 4.0-6.0 and a density of 1.0100 to 1.0300 g/ml.

A watercress extract was obtained from Nasturtium officinal using twovolumes of a solvent containing water, propylene glycol, and ethanol(43%, 42%, and 15%, respectively). The extract thus obtained was solublein water, 60% ethanol, or 94% ethanol. It had a density of 1,000-1.010g/ml and a pH value of 5.0-6.0. Active ingredients in this extractincluded mustard glycosides, gluconasturtiine, etheric oils, andraphanol.

A witch hazel extract was prepared from the leaves of Hamamelisvirginiana also using the just-mentioned solvent containing water,propylene glycol, and ethanol. The resultant extract was a brown-yellowliquid with a characteristic aromatic odor, a sweetish taste, a pH valueof 4.0-5.0, and a density of 1.0100-1.0250 g/ml.

A centella extract was prepared using 1:1 ethanol and water in the samemanner described above.

To prepare a licorice extract, finely cut licorice roots were extractedwith cold water. Ethanol was then added to the extract. The precipitatewas retained by sedimentation and mixed with inorganic acid. Theresulting mixture was filtrated. After neutralizing. The mixture wasdissolved in an ammonium solution and evaporated to dry. The remain wasre-crystallized in acetic acid or ethanol. The licorice extract(containing monoammonium glycyrrhizinate) thus-obtained was a white toyellowish crystal powder with a characteristic odor and sweet taste.

EXAMPLE 2

The coding sequence of the human SCAF gene was cloned into a retroviralexpression vector and placed under the control of a CMV promoter. Asequence encoding a truncated NGFR protein was also cloned into thevector and placed under the control of the retroviral LTR. The NGFRprotein was used as a selective marker. All of the constructs wereidentified by restriction enzyme digestion and confirmed by DNAsequencing analysis. A confirmed vector was transfected into mammaliancells to express the SCAF protein.

Expressed SCAF protein was purified by standard procedure, sterilized bymembrane filtration (pore-size of 0.2 μm), and diluted in PBS to form aSCAF-containing composition. The SCAF and the extracts described inExample 1 above were used to prepare a topical composition according tothe following recipe:

TABLE 1 Ingredients w/w (%) Wheat Germ Oil 2.5 Jojoba Oil 3.5Biosaccharide Gum-1 5 Anti-Wrinkle herbal blend 15 Whitening herbalblend 15 Anti-acne herbal blend 15 Glycerin 6 1,3 Butylene Glycol 5Sorbtiol 5 Polysorbate 85 2 Xanthan Gum 0.5 Methylcellulose 0.25Methylparaben 0.1 Proylparaben 0.1 SDF-1 3000 Unit Purified water Add to100 g

EXAMPLE 3

The safety of a SCAF-containing composition was tested in an animalmodel for allergy and other adverse effects.

Female hairless SKH-1 mice, 6-8 weeks old were used. Such mice have beenwidely used for evaluating the safety of a skin protection agent. Morespecifically, 12 hairless SKH-1 mice were divided into two groups:Control group and Experimental group (6 in each). The mice in theControl group were administered with PBS; and the mice in theExperimental group were treated with the SCAF composition at 20 times ofthe working concentration as described in Table 1 above (3000 U/100 g).The PBS or SCAF composition was smeared onto a circled skin area in theflank of each mouse twice per day for 3 months. During this period, eachmouse was observed for behaviors indicating allergy or toxicity (e.g.,uneasiness and scratching) and changes in the body temperature.

All mice survived the 3-month period and showed no change of behaviorsduring this period. The body temperature of each mouse remained in anormal range. Gross skin examination indicated no obvious abnormalities,such as edema, redness of skin, or neoplasm.

At the end of the 1^(st) or 2^(nd) month, a skin sample was obtainedfrom each mouse by biopsy. At the end of the 3^(rd) month, all mice weresacrificed by euthanasia and a skin sample was obtained from each byautopsy. Each sample was then subjected to Haematoxylin & Eosin stainingand examined under a microscope for any obvious abnormality, such asneoplasm, changes in skin structure and morphology, allergy reaction,and others. No abnormality was found in the epidermis (stratum corneum,granulosum, malpighii, and basale), the dermis, and the subcutaneoustissue. The above results demonstrate that the SCAF-containingcomposition is safe and does not cause allergy or other adverse effectsto the skin.

The above-described SCAF-containing composition was tested on humanvolunteers. Each person was smeared with 0.9% NaCl solution on one arm,and the SCAF-containing composition on the other arm (15 time of theworking concentration) twice a day for 3 weeks. The results are shown inTable 2. The experiment was repeated on another group of humanvolunteers except that a SCAF-containing composition of 10 time of theworking concentration was used twice a day for 2 weeks. The results areshown in Table 3.

TABLE 2 Volunteer Person Age Range Allergy Other Symptoms Male 12 25-580 1* Female 18 22-52 0 0  *One volunteer had a very light scratch on thetested area during the testing period. The skin showed a short termredness, but recovered shortly.

TABLE 3 Volunteer Person Age Range Allergy Other Symptoms Male 8 33-55 00 Female 18 28-48 0 0

The results demonstrate that the SCAF-containing composition does notcause allergy or other adverse effects to human skin.

The above experiment was repeated using the composition described abovein Table 1. No allergy or other adverse effects were found.

Further, the composition was tested for its wrinkle-reducing efficacy.More specifically, 32 volunteer female subjects of 25-55 years old wereadministered with the composition for 30 days. All of them showedvisible reducing in wrinkle.

OTHER EMBODIMENTS

All of the features disclosed in this specification may be combined inany combination. Each feature disclosed in this specification may bereplaced by an alternative feature serving the same, equivalent, orsimilar purpose. Thus, unless expressly stated otherwise, each featuredisclosed is only an example of a generic series of equivalent orsimilar features.

From the above description, one skilled in the art can easily ascertainthe essential characteristics of the present invention, and withoutdeparting from the spirit and scope thereof, can make various changesand modifications of the invention to adapt it to various usages andconditions. Thus, other embodiments are also within the scope of thefollowing claims.

1-8. (canceled)
 9. A method of reducing skin wrinkles, comprisingapplying to a surface of skin in need thereof an effective amount of atopical composition comprising an isolated polypeptide having thesequence of SEQ ID NO:
 1. 10. A method of promoting hair growth,comprising applying to a surface of skin in need thereof an effectiveamount of a topical composition comprising an isolated polypeptidehaving the sequence of SEQ ID NO:
 1. 11-12. (canceled)
 13. An isolatednucleic acid comprising a sequence encoding a polypeptide having theamino acid sequence of SEQ ID NO:
 1. 14. An expression vector comprisinga nucleic acid sequence of claim
 13. 15. A host cell comprising anucleic acid sequence of claim
 13. 16. A method of producing apolypeptide, comprising culturing the host cell of claim 15 in a mediumunder conditions permitting expression of a polypeptide encoded by thenucleic acid, and purifying the polypeptide from the cultured cell orthe medium of the cell.
 17. The method of claim 9, wherein thecomposition further comprises an agent selected from the groupconsisting of an anti-wrinkle herbal extract, a whiting herbal extract,and an anti-acne herbal extract.
 18. The method of claim 17, wherein theanti-wrinkle herbal extract contains a hop extract, a horsetail extract,or both.
 19. The method of claim 17, wherein the whiting herbal extractcontains a Scutellaria baicalensis root extract, a Saxifrage sarmentosaroot extract, or both.
 20. The method of claim 17, wherein the anti-acneherbal extract contains a Watercress extract, a witch hazel extract, acentella extract, a licorice extract, or a combination thereof.
 21. Themethod of claim 17, wherein the composition further contains apolysaccharide.
 22. The method of claim 17, where in the compositioncontains L-fucose, D-galactose, galacturonic acid, or a combinationthereof.
 23. The method of claim 17, wherein the anti-wrinkle herbalextract, the whiting herbal extract, and the anti-acne herbal extractare prepared at the w/w ratio of 3-30%, 3-30%, and 3-30%, respectively.24. The isolated nucleic acid of claim 13, wherein the isolated nucleicacid contains the sequence of SEQ ID NO: 1.